A novel ion conducting route besides the central pore in an inherited mutant of G‐protein‐gated inwardly rectifying K⁺ channel

Abstract

G‐protein‐gated inwardly rectifying K+ (GIRK; Kir3.x) channels play important physiological roles in various organs. Some of the disease‐associated mutations of GIRK channels are known to induce loss of K+ selectivity but their structural changes remain unclear. In this study, we investigated the mechanisms underlying the abnormal ion selectivity of inherited GIRK mutants. By the two‐electrode voltage‐clamp analysis of GIRK mutants heterologously expressed in Xenopus oocytes, we observed that Kir3.2 G156S permeates Li+ better than Rb+, while T154del or L173R of Kir3.2 and T158A of Kir3.4 permeate Rb+ better than Li+, suggesting a unique conformational change in the G156S mutant. Applications of blockers of the selectivity filter (SF) pathway, Ba2+ or Tertiapin‐Q (TPN‐Q), remarkably increased the Li+‐selectivity of Kir3.2 G156S but did not alter those of the other mutants. In single‐channel recordings of Kir3.2 G156S expressed in mouse fibroblasts, two types of events were observed, one attributable to a TPN‐Q‐sensitive K+ current and the second a TPN‐Q‐resistant Li+ current. The results show that a novel Li+‐permeable and blocker‐resistant pathway exists in G156S in addition to the SF pathway. Mutations in the pore helix, S148F and T151A also induced high Li+ permeation. Our results demonstrate that the mechanism underlying the loss of K+ selectivity of Kir3.2 G156S involves formation of a novel ion permeation pathway besides the SF pathway, which allows permeation of various species of cations. Key points G‐protein‐gated inwardly rectifying K+ (GIRK; Kir3.x) channels play important roles in controlling excitation of cells in various organs, such as the brain and the heart. Some of the disease‐associated mutations of GIRK channels are known to induce loss of K+ selectivity but their structural changes remain unclear. In this study, we investigated the mechanisms underlying the abnormal ion selectivity of inherited mutants of Kir3.2 and Kir3.4. Here we show that a novel Na+, Li+‐permeable and blocker‐resistant pathway exists in an inherited mutant, Kir3.2 G156S, in addition to the conventional ion conducting pathway formed by the selectivity filter (SF). Our results demonstrate that the mechanism underlying the loss of K+ selectivity of Kir3.2 G156S involves formation of a novel ion permeation pathway besides the SF pathway, which allows permeation of various species of cations.

Licorice metabolite 18β‐glycyrrhetinic acid activates G protein‐gated inwardly rectifying K+ channels

“…Studies with the K ir 3.2 S148F mutant showed that application of 1‐mM GA did not induce a change in the current amplitude (Figure 2f). Mutation of K ir 3.2 Ser148 to Phe alters its ion selectivity (Chen et al, 2022), suggesting a conformational change in the pore‐forming region. GA may therefore be no longer capable of modulating the activity of this mutant channel because of the overall structural difference.…”

“…Site‐directed mutagenesis of K ir 3.1 and K ir 3.2 mutants was performed as previously described (Chen et al, 2022). Briefly, PfuUltra II Fusion HS DNA Polymerase kit (Agilent Technologies, Santa Clara, CA, USA) and the primers (Table S1) were used to introduce point mutation in K ir 3.x channels by polymerase chain reaction and were confirmed by DNA sequencing.…”

Time-course adaptive changes in hippocampal transcriptome and synaptic function induced by simulated microgravity associated with cognition

“…Studies have revealed that mice exhibit impairments in learning and memory when the Epha6 protein is knocked out ( Savelieva et al, 2008 ; Farini et al, 2020 ). Kcnj6, an essential component of the presynaptic membrane, serves as a key regulatory target for hippocampal neuron excitability and synaptic plasticity ( Alfaro-Ruiz et al, 2021 ; Thompson and Satin, 2021 ; Chen et al, 2022 ). These up-regulated genes contribute to the regulation of synapse structure and function.…”

CL-705G: a novel chemical Kir6.2-specific KATP channel opener

“…Oocytes were isolated from female Xenopus laevis as previously described ( Chen et al, 2022 ). Frogs were purchased from Watanabe Zoushoku (Hyogo, Japan) and housed at 18°C in the animal facility of Wakayama Medical University.…”